Infective endocarditis is a serious infection of the heart with mortality rates in excess of 20%. Endocarditis is thought to occur when bacteria or other microbes gain access to the blood, attach to previously damaged heart valves, and multiply. The oral or viridans streptococci are a leading cause of this illness, and among this group, Streptococcus sanguinis is most common. Prevention of endocarditis relies primarily on antibiotic prophylaxis for at-risk patients prior to dental procedures that are likely to result in bacteremia. However, it is now recognized that most cases of streptococcal endocarditis do not result from such procedures but rather from daily activities that lead to bacteremia, which has resulted in a curtailment in dental antibiotic prophylaxis. The identification of virulence determinants could provide new targets for novel prophylactic measures capable of providing continuous protection for persons at risk. Toward this goal, we have performed multiple screens of S. sanguinis mutants for loss of endocarditis virulence in an animal model. These studies have identified a lipoprotein, SsaB, that is uniquely important for endocarditis virulence and is also a promising target for endocarditis therapy or prevention. SsaB belongs to a family of lipoproteins that function in Mn uptake, oxygen tolerance, and virulence in a number of pathogenic streptococcal species. We have recently demonstrated that SsaB shares these functions. Studies to date suggest that these three functions are causally related, with loss of Mn responsible for oxygen sensitivity, and oxygen sensitivity responsible for loss of virulence. However, this has not been formally demonstrated, nor has it been shown how Mn contributes to either property. We will address these questions using four complementary aims: (1) we will assess the relationship between the Mndependent superoxide dismutase (SodA) and SsaB in oxygen tolerance and virulence;(2) we will examine the effect of Mn limitation on global gene expression;(3) we will examine second-site mutations for their ability to restore oxygen tolerance and virulence to an ssaB mutant;and (4) we will examine the physiological effects of Mn starvation in S. sanguinis. We believe these approaches differ in ways that are critical from previous studies performed with other streptococci. Collectively, these studies will address for the first time the mechanism(s) by which an LraI protein and Mn contribute to oxygen tolerance and virulence in an endocarditis pathogen or, indeed, in any streptococcal pathogen.